BackgroundThe similarities between swine and humans in physiological and genomic patterns, and the great correlation in size and anatomy, make pigs extremely useful in preclinical studies. New-born piglets can represent a model for congenital and genetic diseases in new-born children. It is known that piglets may have significant differences in clinicopathological results compared to adult pigs. Therefore, adult laboratory reference intervals cannot be applied to piglets. The aim of this study was to compare haematological and chemical variables in piglets of two ages and determinate age-related reference intervals for commercial hybrid young pigs.Blood samples were collected under general anaesthesia from 130 animals divided into five- (P5) and 30- (P30) day-old piglets. Only P30 animals were treated with parenteral iron after birth. Samples were analysed using automated haematology (ADVIA 2120) and chemistry analysers, and age-related reference intervals were calculated.ResultsSignificant higher values of RBC, Hb and HCT were observed in P30 animals when compared to P5, with an opposite trend for MCV. These results were associated with a reduction of the RBC regeneration process and the thrombopoietic response. The TSAT and TIBC were significantly higher in P30 compared to P5; however, piglets remained iron deficient compared to adult reference intervals reported previously.ConclusionsIn conclusion, this paper emphasises the high variability occurring in clinicopathological variables between new-born and 30-day-old pigs, and between piglets and adult pigs. This study provides valuable reference data for piglets at precise ages and could be used in the future as historical control improving the Reduction in animal experiments, as suggested by the 3Rs principle.Electronic supplementary materialThe online version of this article (doi:10.1186/s12917-017-0946-2) contains supplementary material, which is available to authorized users.

Essential oils possess a variety of biological properties (i.e., antioxidant, antibacterial, and cytotoxic) that could possibly be applied in reproductive medicine, but their effects on spermatozoa are still partially unknown. The aim of the study was to describe the effects of Thymbra capitata (L.) Cav. and Rosmarinus officinalis (L.) essential oils on the main morpho-functional parameters of swine spermatozoa. Essential oils were preliminary characterized by gas chromatography and added with emulsifiers to facilitate diffusion. Experimental samples were prepared by suspending a fixed number of spermatozoa in 5 mL of medium with 10 different concentrations of essential oil (0.2–2 mg/mL, at intervals of 0.2). After 3 h of incubation, samples were analyzed for pH, viability, objective motility, and acrosome status. Results showed that the effects of the essential oils are concentration-dependent and that R. officinalis is well tolerated up to 0.6 mg/mL. T. capitata impaired the spermatozoa starting from the lowest concentration, with complete spermicidal effect from 0.4 mg/mL. The patterns of damage, confirmed by SEM, were different and quite distinct. As expected, spermatozoa proved to be sensitive to external stimuli and capable of showing different functional patterns, providing interesting insights to the action/toxicity mechanisms. The results of the present work represent the first step towards the systematic characterization of the effects of these compounds on spermatozoa. This kind of studies are necessary to strengthen the idea of future applications of essential oils in the reproductive field due to their antioxidant, antibacterial, or spermicidal properties.

The growing interest towards essential oils stems from their biological capabilities that include antibacterial and antioxidant effects. Such properties may be extremely useful in the reproductive field; nonetheless essential oils show toxic effects that can lead to cell disruption. The present study aimed to evaluate and compare the effects of tea tree oil (TTO) and its principal component terpinen-4-ol (TER) on the morpho-functional parameters of swine spermatozoa. Experimental samples were prepared by suspending 15 × 107 spermatozoa in 5 mL of medium with different concentrations of the above-mentioned compounds: from 0.2 to 2 mg/mL at an interval of 0.2 for TTO, while TER concentrations were adjusted according to its presence in TTO (41.5%). After 3 h incubation at 16 °C, samples were analyzed for pH, viability, acrosome status, and objective motility. The results highlighted a concentration-dependent effect of TTO with total motility as the most sensitive parameter. TER was better tolerated, and the most sensitive parameters were related to membrane integrity, suggesting a different pattern of interaction. The study confirms the importance of evaluating the effects of natural compounds on spermatozoa before exploiting their beneficial effects. Spermatozoa seem to be good candidates for preliminary toxicological screenings in the light of their peculiar properties.

When it comes to neuroscience, pigs represent an important animal model due to their resemblance with humans’ brains for several patterns including anatomy and developmental stages. Cerebrospinal fluid (CSF) is a relatively easy-to-collect specimen that can provide important information about neurological health and function, proving its importance as both a diagnostic and biomedical monitoring tool. Consequently, it would be of high scientific interest and value to obtain more standard physiological information regarding its composition and dynamics for both swine pathology and the refinement of experimental protocols. Recently, proton nuclear magnetic resonance (1H NMR) spectroscopy has been applied in order to analyze the metabolomic profile of this biological fluid, and results showed the technique to be highly reproducible and reliable. The aim of the present study was to investigate in both qualitative and quantitative manner the composition of Cerebrospinal Fluid harvested form healthy newborn (5 days old-P5) and young (30-P30 and 50-P50 days old) piglets using 1H NMR Spectroscopy, and to analyze any possible difference in metabolites concentration between age groups, related to age and Blood-Brain-Barrier maturation. On each of the analyzed samples, 30 molecules could be observed above their limit of quantification, accounting for 95–98% of the total area of the spectra. The concentrations of adenine, tyrosine, leucine, valine, 3-hydroxyvalerate, 3-methyl-2-oxovalerate were found to decrease between P05 and P50, while the concentrations of glutamine, creatinine, methanol, trimethylamine and myo-inositol were found to increase. The P05-P30 comparison was also significant for glutamine, creatinine, adenine, tyrosine, leucine, valine, 3-hydroxyisovalerate, 3-methyl-2-oxovalerate, while for the P30-P50 comparison we found significant differences for glutamine, myo-inositol, leucine and trimethylamine. None of these molecules showed at P30 concentrations outside the P05 –P50 range.

Simple SummaryThe roe deer is a very common wild species in Italy and shows peculiar reproductive characteristics. Sexually-mature males, called bucks, show a complete interruption in spermatogenesis during the cold seasons. The mechanisms behind such interruption are still partially unknown. Hair is a good biological sample, easy to obtain while minimizing stress, for endocrinological analyses that may provide information regarding such mechanisms. The aim of the work was to quantify and compare hair concentrations of testosterone and cortisol in wild roe deer bucks hunted during the pre- and post-rut period (mating season). The secondary objective was the evaluation of possible correlations of such hair concentrations with blood and morphometric parameters of the testes. Testosterone significantly increased from the pre- to post-rut period, while cortisol significantly decreased. The correlations with blood and testicular parameters resemble what is already described in the literature. Overall, this study represents a first report of the quantification of testosterone and cortisol in roe deer hair, and may provide interesting insights into their reproductive physiology.AbstractThe roe deer is a seasonally breeding species with a reproductive cycle regulated by endogenous rhythms and photoperiod-sensitivity. Sexually mature bucks show hormonal and testicular activation during the reproductive season, with a peak in the rut period, and following gradual involution. Hair is a good matrix for non-invasive endocrinological analyses that provide long-term information without being influenced by the hormones’ pulsating release patterns in blood. The aim of the work was to quantify hair concentrations of testosterone and cortisol in wild roe deer bucks hunted during the pre- and post-rut period, using a radioimmunoassay methodology, and to look for differences between the two periods. The secondary objective was the evaluation of possible correlations of such hair concentrations with blood and morphometric parameters of the testes. Both hormones showed statistical differences, with opposing trends, when comparing the two periods: testosterone increased while cortisol decreased. The correlation analysis was in agreement with existing literature regarding metabolism/actions of these hormones and testicular morphometric parameters. This study represents the first report of the use of radioimmunoassay techniques to quantify testosterone and cortisol in roe deer hair, and may provide interesting insights into their reproductive physiology.

Antimicrobial resistance is increasing within the porcine industry with consequential high impact on human health, leading to a need for new antimicrobials. Lately, the scientific community has turned its interest towards natural compounds, and different essential oils have been tested on spermatozoa for preliminary assessment of toxicity before considering them as good substitutes for standard antibiotics. The aim of the present work was to investigate the potential antimicrobial effect of Melaleuca alternifolia and Rosmarinus officinalis essential oils, already evaluated for toxicity, on swine artificial insemination doses deprived of spermatozoa and stored at 16 °C for 5 days. This was accomplished by setting up an in vitro model with a standardized quantity of E. coli. Essential oils, previously chemo-characterized by means of gas chromatography, were tested at 0.2 and 0.4 mg/ml.Analyses, performed at 24 and 120 hours, included optical density evaluation, bacterial DNA quantification by qPCR, and colony count. The results demonstrate that both Melaleuca alternifolia and Rosmarinus officinalis essential oils, at a concentration of 0.4 mg/ml, are capable of delivering similar effects to ampicillin, used as control, on the experimental samples. At the lower concentration, M. alternifolia essential oil seemed more effective when compared to R. officinalis. Overall, these findings strengthen the hypothesis of the potential use of phyto-complexes as antimicrobial agents for reproductive biotechnologies.

Mucopolysaccharidosis type IIIA (MPS-IIIA) is a lysosomal storage disorder (LSD) caused by inherited defect of sulfamidase, a lysosomal sulfatase. MPS-IIIA is one of the most common and severe forms of LSDs with CNS involvement. Presently there is no cure. Here we have developed a new gene delivery approach for the treatment of MPS-IIIA based on the use of a modified version of sulfamidase expression cassette. This cassette encodes both a chimeric sulfamidase containing an alternative signal peptide (sp) to improve enzyme secretion and sulfatase-modifying factor 1 (SUMF1) to increase sulfamidase post-translational activation rate. We demonstrate that improved secretion and increased activation of sulfamidase act synergistically to enhance enzyme biodistribution in wild-type (WT) pigs upon intrathecal adeno-associated virus serotype 9 (AAV9)-mediated gene delivery. Translating such gene delivery strategy to a mouse model of MPS-IIIA results in a rescue of brain pathology, including memory deficit, as well as improvement in somatic tissues. These data may pave the way for developing effective gene delivery replacement protocols for the treatment of MPS-IIIA patients.

Vascularization is a crucial factor when approaching any engineered tissue. Vascular wall-mesenchymal stem cells are an excellent in vitro model to study vascular remodeling due to their strong angiogenic attitude. This study aimed to demonstrate the angiogenic potential of experimental highly porous scaffolds based on polylactic acid (PLA) or poly-e-caprolactone (PCL) doped with calcium silicates (CaSi) and dicalcium phosphate dihydrate (DCPD), namely PLA-10CaSi-10DCPD and PCL-10CaSi-10DCPD, designed for the regeneration of bone defects. Vascular wall-mesenchymal stem cells (VW-MSCs) derived from pig thoracic aorta were seeded on the scaffolds and the expression of angiogenic markers, i.e. CD90 (mesenchymal stem/stromal cell surface marker), pericyte genes α-SMA (alpha smooth muscle actin), PDGFR-β (platelet-derived growth factor receptor-β), and NG2 (neuron-glial antigen 2) was evaluated. Pure PLA and pure PCL scaffolds and cell culture plastic were used as controls (3D in vitro model vs. 2D in vitro model). The results clearly demonstrated that the vascular wall mesenchymal cells colonized the scaffolds and were metabolically active. Cells, grown in these 3D systems, showed the typical gene expression profile they have in control 2D culture, although with some main quantitative differences. DNA staining and immunofluorescence assay for alpha-tubulin confirmed a cellular presence on both scaffolds. However, VW-MSCs cultured on PLA-10CaSi-10DCPD showed an individual cells growth, whilst on PCL-10CaSi-10DCPD scaffolds VW-MSCs grew in spherical clusters. In conclusion, vascular wall mesenchymal stem cells demonstrated the ability to colonize PLA and PCL scaffolds doped with CaSi-DCPD for new vessels formation and a potential for tissue regeneration.